Target (protein/gene name): Hypoxanthine guanine phophoribosyltransferase (HGPTR) NCBI Gene # or RefSeq#: NC_000962.3 Protein ID (NP or XP #) or Wolbachia#: NP_218141 Organism (including strain): Mycobacterium tuberculosis (H37Rv) Etiologic Risk Group (see link below): N/A Disease Information (sort of like the Intro to your Mini Research Write up): Mycobacterium tuberculosis is a pathogenic bacteria responsible for the infectious disease called Tuberculosis. Tuberculosis is highly virulent, having a high morbidity and mortality rate compared to other diseases. The disease began increasing after 1985 when HIV infections were introduced that lowered the immunity of the body. The virulence of the bacteria is transferred by microscopic airborne bodily fluids that infect the alveolar surfaces in the lungs. Some common symptoms include coughing, chest pain, fatigue, fever and etc. People who live in areas that lack medical care, have poor nutritional resources, and are overcrowded risk exposure to Tuberculosis. Each year, mutations have lead to more resistant strains of M. tuberculosis, so research is always ongoing to counteract the spreading of the disease. Link to TDR Targets page (if present): http://www.tdrtargets.org/targets/view?gene_id=6224 Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.) https://www.patricbrc.org/view/Feature/PATRIC.83332.12.NC_000962.CDS.4063254.4063856.rev#view_tab=overview https://www.ncbi.nlm.nih.gov/gene/?term=2.4.2.8+mycobacterium+tuberculosis
Essentiality of this protein: Is it a monomer or multimer as biological unit?: The most probable structure of HGPTR is a dimer.
HGPTR plays a role in nucleotide salvage pathways. This protein is essential in M. tuberculosis and other organisms for the recycling of the purine nucleotide for DNA replication and transcription. Salvaging nucleotides will allow the infectious organism to replicate its genetic material efficiently to be carried out in a viral pathway in the host. There were also essentiality data on TDR targets that described HGPTR as gaining significant fitness.
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism): [1] Eng, W.S.; Hockova, D.; Spacek P.; Janeba, Z.; West, N.P.; Woods, K.; Naesens, L.M.J.; Keough, D.T.; Guddat, L.W. First Crystal Structures of Mycobacterium tuberculosis 6-Oxopurine Phosphoribosyltransferase: Complexes with GMP and Pyrophosphate and with Acyclic Nucleoside Phosphonates Whose Prodrugs Have Antituberculosis Activity. Journal of Medicinal Chemistry2015, 58, (11), 4822-4838
Structure (PDB or Homology model) -- PDB # or closest PDB entry if using homology model: 5KNP
Current Inhibitors: N/A in M. tuberculosis Expression Information (has it been expressed in bacterial cells):
Biazus, G.; Schneider, C.Z.; Palma, M.S.; Basso, L.A.; Santos, D.S. Hypoxanthine-guanine phosphoribosyltransferase from Mycobacterium tuberculosis H37Rv: Cloning, expression, and biochemical characterization. Protein Expression and Purification2009, 66, (2), 185-190
Image of protein (PyMol with features delineated and shown separately):
Fig 1. Crystal structure of Mycobacterium tuberculosis hypoxanthine guanine phophoribosyltransferase (5KNP) in complex with [3S,4R]-(4-(Hypoxanthin-9-yl)pyrrolidin-3-yl)-oxymethanephosphonic acid
Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
length of your protein in Amino Acids
207 AA Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website 22.94284 kDa Molar Extinction coefficient of your protein at 280 nm wavelength: 18910 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it. Fig 2. TMpred output for hypoxanthine guanine phophoribosyltransferase
Do Not Need this info for Spring (but still copy these lines to your Target page for now) Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers): (link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol) -- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences): **
NCBI Gene # or RefSeq#: NC_000962.3
Protein ID (NP or XP #) or Wolbachia#: NP_218141
Organism (including strain): Mycobacterium tuberculosis (H37Rv)
Etiologic Risk Group (see link below): N/A
Disease Information (sort of like the Intro to your Mini Research Write up): Mycobacterium tuberculosis is a pathogenic bacteria responsible for the infectious disease called Tuberculosis. Tuberculosis is highly virulent, having a high morbidity and mortality rate compared to other diseases. The disease began increasing after 1985 when HIV infections were introduced that lowered the immunity of the body. The virulence of the bacteria is transferred by microscopic airborne bodily fluids that infect the alveolar surfaces in the lungs. Some common symptoms include coughing, chest pain, fatigue, fever and etc. People who live in areas that lack medical care, have poor nutritional resources, and are overcrowded risk exposure to Tuberculosis. Each year, mutations have lead to more resistant strains of M. tuberculosis, so research is always ongoing to counteract the spreading of the disease.
Link to TDR Targets page (if present):
http://www.tdrtargets.org/targets/view?gene_id=6224
Link to Gene Database page (NCBI, EuPath databases -e.g. TryTryp, PlasmoDB, etc - or PATRIC, etc.)
https://www.patricbrc.org/view/Feature/PATRIC.83332.12.NC_000962.CDS.4063254.4063856.rev#view_tab=overview
https://www.ncbi.nlm.nih.gov/gene/?term=2.4.2.8+mycobacterium+tuberculosis
Essentiality of this protein:
Is it a monomer or multimer as biological unit?: The most probable structure of HGPTR is a dimer.
HGPTR plays a role in nucleotide salvage pathways. This protein is essential in M. tuberculosis and other organisms for the recycling of the purine nucleotide for DNA replication and transcription. Salvaging nucleotides will allow the infectious organism to replicate its genetic material efficiently to be carried out in a viral pathway in the host. There were also essentiality data on TDR targets that described HGPTR as gaining significant fitness.
Druggable Target (list number or cite evidence from a paper/database showing druggable in another organism):
[1] Eng, W.S.; Hockova, D.; Spacek P.; Janeba, Z.; West, N.P.; Woods, K.; Naesens, L.M.J.; Keough, D.T.; Guddat, L.W. First Crystal Structures of Mycobacterium tuberculosis 6-Oxopurine Phosphoribosyltransferase: Complexes with GMP and Pyrophosphate and with Acyclic Nucleoside Phosphonates Whose Prodrugs Have Antituberculosis Activity. Journal of Medicinal Chemistry 2015, 58, (11), 4822-4838
[2] http://www.bindingdb.org/jsp/dbsearch/PrimarySearch_ki.jsp?tag=pol&submit=Search&target=hypoxanthine-guanine%20phosphoribosyltransferase&polymerid=50002984
EC#: 2.4.2.8
Link to BRENDA EC# page:
http://www.brenda-enzymes.org/enzyme.php?ecno=2.4.2.8&Suchword=&reference=&UniProtAcc=&organism%5B%5D=Mycobacterium+tuberculosis&show_tm=0
Link to BRENDA HPTR Pathway:
http://www.brenda-enzymes.org/pathway_index.php?menu=off&site=pathway&search_mode=exact&pathway=purine_metabolism&ecno=2.4.2.8
Enzyme Assay information (spectrophotometric, coupled assay ?, reagents):
https://www.sigmaaldrich.com/content/dam/sigma-aldrich/docs/Sigma/Enzyme_Assay/hypoxanthineguanine.pdf.
Structure (PDB or Homology model)
-- PDB # or closest PDB entry if using homology model:
5KNP
Current Inhibitors: N/A in M. tuberculosis
Expression Information (has it been expressed in bacterial cells):
Biazus, G.; Schneider, C.Z.; Palma, M.S.; Basso, L.A.; Santos, D.S. Hypoxanthine-guanine phosphoribosyltransferase from Mycobacterium tuberculosis H37Rv: Cloning, expression, and biochemical characterization. Protein Expression and Purification 2009, 66, (2), 185-190
Image of protein (PyMol with features delineated and shown separately):
Fig 1. Crystal structure of Mycobacterium tuberculosis hypoxanthine guanine phophoribosyltransferase (5KNP) in complex with [3S,4R]-(4-(Hypoxanthin-9-yl)pyrrolidin-3-yl)-oxymethanephosphonic acid
Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
length of your protein in Amino Acids
207 AA
Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website
22.94284 kDa
Molar Extinction coefficient of your protein at 280 nm wavelength:
18910
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
Fig 2. TMpred output for hypoxanthine guanine phophoribosyltransferase
CDS Gene Sequence (paste as text only):
Protein Sequence (paste as text only):
Do Not Need this info for Spring (but still copy these lines to your Target page for now)
Primer design results for pNIC-Bsa4 cloning (list seqeunces of all of your ~40 nt long primers):
(link to DNA Works output text file - that should be saved in your Google Docs folder after you did the primer design protocol)
-- Ask a mentor, Dr. B, or a fellow researcher -how to link a GDocs file if you are not sure how to.
Primer design results for 'tail' primers (this is just 2 sequences):
**